The methodology of horseradish peroxidase neurohistochemistry will be investigated. Although this procedure is very extensively used, its application to the tracing of cerebral connection often suffers from serious methodological shortcomings which jeopardize the validity of many of the results. In this application, we propose to investigate the determinants of such methodological pitfalls and to suggest new procedures for circumventing these. We will concentrate on three areas: 1) The improvement of sensitivity so that the full extent of transported HRP, anterograde as well as retrograde, may be visualized with greater acuity than is presently possible. 2) The improvement of reliability by investigating the effective injection site, the contribution of passing fibers and the spurious labeling of neurons. 3) The development of a non-carcinogenic chromogen to eliminate a laboratory health hazard. With this methodology, we will study issues in cerebral connectivity of the monkey brain which are related to our work in the past and which require this type of methodology for definitive analysis. These areas are: 1) The neural afferents of association cortex. 2) The neocortical input into limbic structures such as the nucleus accumbens septi, septal area, hippocampus and olfactory bulb. 3) The functional and topographical organization of pulvinar efferents to neocortex. 4) The histochemical characterization of the cholinergic connections from basal forebrain to neocortex and from septum to the hippocamus. In these studies, amino acid autoradiography, acetylocholinesterase and choline acetylase histochemistry and QNB autoradiography will also be used. While our methodological work will improve the scope and power of HRP histochemistry, the connectivistic investigations will provide information which is presently not available and which is closely related to issues of cognition, memory and dementia in the human.